Peroxynitrite (ONOO ), formed from the reaction between superoxide and nitric oxide, is a potent oxidant that can oxidize a variety of biomolecules such as glutathione (GSH), lipids, and DNA. The exact molecular mechanism of oxidation in these systems has not been established; however, free radical intermediacy has been proposed. The one-electron oxidation of biologically-relevant compounds (e.g. `-tocopherol, ascorbic acid, methionine) by ONOO in defined chemical systems is, however, a minor pathway and it appears that the major pathway involves two-electron oxidation. Thus, the toxicity of peroxynitrite in biological systems is unlikely to involve one electron mediated-oxidation of antioxidants such as `-tocopherol and ascorbic acid. The ESR spin trapping is the most suitable physical technique for detecting and characterizing thiyl radical reactions at room temperature. Peroxynitrite-mediated oxidation of thiols such as glutathione, cysteine, and N-acetyl-dl-penicillamine in the presence of DMPO produced the corresponding DMPO-thiyl radical adducts. The DMPO/ OH adduct formed in these systems was inhibited in the presence of DMPO/ OOH. It is likely that reduction of DMPO/ OOH to DMPO/ OH is enhanced by thiols. The spectral intensity of DMPO-thiyl adducts was enhanced by SOD and SOD mimic, indicating a superoxide-mediated destruction of DMPO-thiyl radical adducts. Peroxynitrite oxidized formate to CO2 and CO2 in the presence of thiols. Based on photolysis experiments of nitrosothiols, we conclude that GS and other thiyl radicals are responsible for oxidation of formate to CO2 [RS + HCO2 RSH + CO2 ]. Peroxynitrite-mediated oxidation of sulfite in the presence of DMPO produced DMPO/ SO3 adduct. In the presence of formate, the DMPO/ CO2 adduct was observed, suggesting that an oxidant derived from SO3 and O2 is responsible for oxidation of formate to CO2 . The reduction between peroxynitrite and GSH can amplify the oxidative damage via formation of O2 , which may limit the antioxidant potential of GSH. Thus, addition of SOD is required to inhibit peroxynitrite-mediated superoxide formation.